Biologically derived fuels are viable alternatives to traditional fossil fuels, and microalgae are a particularly promising source, but improvements are required throughout the production process to increase productivity and reduce cost. Metabolic engineering to increase yields of biofuel-relevant lipids in these organisms without compromising growth is an important aspect of advancing economic feasibility. We report that the targeted knockdown of a multifunctional lipase/phospholipase/acyltransferase increased lipid yields without affecting growth in the diatom Thalassiosira pseudonana. Antisense-expressing knockdown strains 1A6 and 1B1 exhibited wild-type-like growth and increased lipid content under both continuous light and alternating light/dark conditions. Strains 1A6 and 1B1, respectively, contained 2.4-and 3.3-fold higher lipid content than wild-type during exponential growth, and 4.1-and 3.2-fold higher lipid content than wild-type after 40 h of silicon starvation. Analyses of fatty acids, lipid classes, and membrane stability in the transgenic strains suggest a role for this enzyme in membrane lipid turnover and lipid homeostasis. These results demonstrate that targeted metabolic manipulations can be used to increase lipid accumulation in eukaryotic microalgae without compromising growth. metabolism | RNAi | algal biofuel | targeted manipulation | triacylglycerol

Coral bleaching is the breakdown of symbiosis between coral animal hosts and their dinoflagellate algae symbionts in response to environmental stress. On large spatial scales, heat stress is the most common factor causing bleaching, which is predicted to increase in frequency and severity as the climate warms. There is evidence that the temperature threshold at which bleaching occurs varies with local environmental conditions and background climate conditions. We investigated the influence of past temperature variability on coral susceptibility to bleaching, using the natural gradient in peak temperature variability in the Gilbert Islands, Republic of Kiribati. The spatial pattern in skeletal growth rates and partial mortality scars found in massive Porites sp. across the central and northern islands suggests that corals subject to larger year-to-year fluctuations in maximum ocean temperature were more resistant to a 2004 warm-water event. In addition, a subsequent 2009 warm event had a disproportionately larger impact on those corals from the island with lower historical heat stress, as indicated by lower concentrations of triacylglycerol, a lipid utilized for energy, as well as thinner tissue in those corals. This study indicates that coral reefs in locations with more frequent warm events may be more resilient to future warming, and protection measures may be more effective in these regions.

Depth is used often as a proxy for gradients in energetic resources on coral reefs and for predicting patterns of community energy use. With increasing depth, loss of light can lead to a reduced reliance on autotrophy and an increased reliance on heterotrophy by mixotrophic corals. However, the generality of such trophic zonation varies across contexts. By combining highresolution oceanographic measurements with isotopic analyses (d 13 C, d 15 N) of multiple producer and consumer levels across depths (10-30 m) at a central Pacific oceanic atoll, we show trophic zonation in mixotrophic corals can be both present and absent within the same reef system. Deep-water internal waves that deliver cool particulate-rich waters to shallow reefs occurred across all sites (2.5-5.6 events week-1 at 30 m) but the majority of events remained depthrestricted (4.3-9.7 % recorded at 30 m propagated to 10 m). In the absence of other particulate delivery, mixotrophs increased their relative degree of heterotrophy with increasing depth. However, where relatively long-lasting downwelling events (1.4-3.3 times the duration of any other site) occurred simultaneously, mixotrophs displayed elevated and consistent degrees of heterotrophy regardless of depth. Importantly, these long-lasting surface pulses were of a lagoonal origin, an area of rich heterotrophic resource supply. Under such circumstances, we hypothesize heterotrophic resource abundance loses its direct linkage with depth and, with resources readily available at all depths, trophic zonation is no longer present. Our results suggest that fine-scale intra-island hydrographic regimes and hydrodynamic connectivity between reef habitats contribute to explaining the context specific nature of coral trophic depth zonation in shallow reef ecosystems.

Mutually beneficial interactions between species (mutualisms) shaped the evolution of eukaryotes and remain critical to the survival of species globally [1, 2]. Theory predicts that hosts should pass mutualist symbionts to their offspring (vertical transmission) [3-8]. However, offspring acquire symbionts from the environment in a surprising number of species (horizontal acquisition) [9-12]. A classic example of this paradox is the reef-building corals, in which 71% of species horizontally acquire algal endosymbionts [9]. An untested hypothesis explaining this paradox suggests that horizontal acquisition allows offspring to avoid symbiont-induced harm early in life. We reconstructed the evolution of symbiont transmission across 252 coral species and detected evolutionary transitions consistent with costs of vertical transmission among broadcast spawners, whose eggs tend to be positively buoyant and aggregate at the sea surface. Broadcasters with vertical transmission produce eggs with traits that favor reduced buoyancy (less wax ester lipid) and rapid development to the swimming stage (small egg size), both of which decrease the amount of time offspring spend at the sea surface. Wax ester provisioning decreased after vertically transmitting species evolved brooding from broadcasting, indicating that reduced buoyancy evolves only when offspring bear symbionts. We conclude that horizontal acquisition protects offspring from damage caused by high light and temperatures near the sea surface while providing benefits from enhanced fertilization and outcrossing. These findings help explain why modes of symbiont transmission and reproduction are strongly associated in corals and highlight benefits of delaying mutualist partnerships, offering an additional hypothesis for the pervasiveness of this theoretically paradoxical strategy.